Although about 40% of the world population is at risk of deadly parasitic disease infections, there are insufficient safe, reliable drugs for treatment of or under development for these diseases. The field is limited by ideas for novel cellular receptors or types of drugs. The research in this proposal is focused on methods to address both problems. We propose compounds that can selectively target a unique cellular target, the thousands of AT-rich, DNA minicircles that are interlocked into the parasite mitochondrial kinetoplast genome. Our proposal has plans for innovative approaches to inhibit the complex replication reactions of the minicircles involved with opening, copying and restructuring daughter/parent kinetoplasts. We will approach the problem from a fundamental basis and will design and synthesize new types of compounds to interfere with kinetoplast replication. We will conduct biophysical studies on both model and kinetoplast DNAs with the innovative new compounds and the results will be correlated with cell uptake and distribution studies that are done by collaborating groups of recognized parasite biologists. Three specific aims describe new directions in our research that are largely based on discoveries from the funded project. Our general hypothesis is: we can establish a fundamental basis for the design of new types of compounds that have therapeutic potential as a result of synergistic effects on the nonstandard DNA sequences and structures of the kinetoplast. To do this research two collaborating groups will conduct focused compound synthesis along with biophysical characterization of DNA complexes to answer specific questions that are very difficult to answer by other approaches. Under aim 1 we build on a discovery that shows the classical model for minor groove binding is too limited and that linear compounds can bind strongly and specifically to DNA by using interfacial water. We will explore the limits on linear compound binding and determine if there is a thermodynamic signature for complexes with a bound water. Under aim 2 we propose completely new types of compounds, which are designed to mimic protein motifs and cause significant bending of DNA. One set uses two connected AT site binding units with a short linker to bend the helix into the minor groove. The other set uses a strong binding minor groove motif with a partial intercalating wedge to bend DNA into the major groove. Such effects on structure should be particularly pronounced at the kinetoplast of parasites. Under aim 3 we use the fact that kinetoplasts are AT rich but their AT sequences are broken into small units that are typically separated by one or two GC base pairs. We propose compounds with strong-binding AT motifs that are linked with groups that specifically recognize intervening GC base pairs. This added GC selectivity, coupled to specific terminal AT recognizing motifs, will provide high specificity for sites that are quite common in kinetoplast DNA. We have a unique, collaborative research team, which has rewritten the mechanism for small molecule-minor groove complex formation and for design of compounds for DNA therapeutics, to carry out this research.